Efficiency of eugenol oil nanoemulsion against Banana bunchy top virus and contamination with fungi in plant tissue culture

Development of micropropagation technique for mass propagation of virus-free banana 
plantlets and reduction of the frequency of fungal contamination is critical. For this purpose the 
experimental trials in the current study aimed to evaluate the effectiveness of eugenol oil 
nanoemulsion (EON) on elimination of Banana bunchy top virus (BBTV) and fungal contaminants 
of banana under in vitro conditions. To achieve our aims, the virus has been identified on the basis 
of symptoms from naturally infected banana seedlings with its suckers (Musa spp. cv. Grand Nain), 
grown in Qalyubia Governorate, and later by serological detection (DAS-ELISA) as BBTV then 
used as a source for virus-infected explants in a tissue culture line. Three fungal contaminants 
(Aspergillus flavus, Aspergillus niger and Penicillium expansum) were identified in growth media of 
banana in vitro cultures on the basis of colony morphology and microscopic appearance. The rate 
of occurrence of aspergilli (Aspergillus flavus and A. niger) was found to be higher than Penicillium 
expansum. Different concentrations (125, 250 and 500 mg/L) from EON were studied in vitro. All 
tested concentrations exhibited high inhibitory activities (94.81, 95.5and 96.66 %) against 
Penicillium expansum and almost the same effect (88%) on Aspergillus niger but fluctuated their 
effect on A. flavus causing a decrease in the size and rate of growth by 79.99, 80.73 and 77.77%, 
respectively. On the other side, the inhibition rate of virus activity caused by treatments was 
evaluated using DAS-ELISA. Results showed that there was no antiviral effect of EON at 
concentration 125 mg/L compared with concentrations of 250 and 500 mg/L of EON which caused 
a very low antiviral activity through primary establishment stage. The antiviral effect of EON seems 
to be based on the incorporation of EON particles, three successive times, into culture MS media 
during implementation of the in vitro multiplication. Preliminary results indicate a possibility of 
therapeutically application of EON at high concentration (500 mg/L) against BBTV with completely 
decontamination of culture medium in plant tissue culture technique. Moreover, root induction was 
achieved on MS medium containing 1.0 mg/L indole-3-butyric acid and complete BBTV-free banana 
plantlets were obtained after five weeks in vitro.  
Keywords: Banana, Banana bunchy top virus, M